The Effect Of Ethanol Stress On The Metabolic Activity And Membrane Structure Of Lactobacillus

Lactic acid bacteria(LAB) are a kind of microbes which are produced lactic acid. It is important starter cultures and popular probiotics in the fermentation industry. However, the accumulation of ethanol during the interaction and development between the LAB and yeasts, and becoming an” ethanol stress” environment. It could cause metabolic of bacteria disorder, and even death. The primary issue is the way to improve the vitality of LAB by ethanol stress in the food fermentation industry. In this dissertation, taking Lactobacillus as experimental object, the physiological changes of bacteria and its damage mechanisms have been examined dur ing ethanol stress process. Meanwhile an ethanol-resistant adaptive strain was obtained by adaptive domestication, and damage mechanisms were further discussed in the paper.In order to analyze the main factors causing the loss of the survival rates of bacteria, in this study, the survival rates of bacteria was investigated during different treatments concentrations of ethanol. In addition, the activity changes of hexokinase(HK), pyruvate kinase(P K),lactate dehydrogenase(LDH), ATPases and changes of cell membrane with stress treatments w ere determined relatively.Taking Lactobacillus crustoru m D2-5, Lactobacillus plantarum D5-5 and Lactobacillus casei D5-8 as experimental objects, the experiment compared their activities change after ethanol stress. The experiment results demonstrated that the Lactobacillus growth dropped while the activities reduced on the contrary with the increase of the ethanol concentration. The survival rate has the maximum decline with 8%(v/v) ethanol concentration of stress, Lactobacillus crustorum D2-5, Lactobacillus plantarum D5-5 and Lactobacillus casei D5-8 survival rate of only 26.6%, 23.5% and 34.2%. It also suggested that the differences in HK during different ethanol concentration treatments were not significant but the activities of PK, LDH and ATPase were significant, the extracellular β-galactosidase activity was increased. Meanwhile, the membrane integrity was decreased and cell ultrastructure was injured in the presence of ethanol stress treatment. These results indicated that PK, LDH and ATPase are the key factors of damage caused by ethanol stress and could be used to evaluate the effect of ethanol stress on viability and metabolic activities of Lactobacillus, the activity changes associated with strains viability.Taking Lactobacillus plantarum D5-5 as the main study objects, to examine its damage mechanisms during ethanol stress process, in this paper, we investigated the activity of bacteria and changes of key enzymes and membrane permeability. The results showed that the survival rate of Lactobacillus plantarum D5-5 was decreased by stress treatment, but the p H was increased. The survival rate has the maximum decline with 8%(v/v) ethanol concentration of stress, and decreased by 69.79%, p H improved by 1.58. The activities of LDH and ATPase were significantly different prior to and after stressing, but extracellular β-galactosidase activity was increased. The above results indicated that the key enzymes activity loss and the membrane permeability were changed by stress treatments. These results have provided direct viability and metabolic evidence of injured cell with stressing.In this dissertation, Lactobacillus crustorum D2-5, Lactobacillus plantarum D5-5 and Lactobacillus casei D5-8 as an initial strain, and an ethanol-resistant adaptive strain was acquired by adaptive domestication. To compared the viability and metabolic changes of initial and adaptive strains during ethanol stress. We determined the survival of evolved bacteria, vitality, key enzymes, membrane integrity and permeability. The results showed that the survival rate of the adaptive strain was increased by stress treatment. Lactobacillus crustorum D2-5 and Lactobacillus plantarum D5-5 survival rates increased by 21.15%、23.21% respectively. The key enzymes activity was significantly improved. The PK, LDH and ATPase were affected signif icantly. This result implied the PK, LDH and ATPase were the key factors of damage caused by stress treatment. The extracellular β-galactosidase activity of adaptive strain was less after stressing. This study indicated the permeability and integr ity of cell membrane was maintained by adaptive domestication in the presence of ethanol stress treatment.